C-terminal signal sequence promotes virulence factor secretion in Mycobacterium tuberculosis

Mycobacterium tuberculosis uses the ESX-1/Snm system [early secreted antigen 6 kilodaltons (ESAT-6) system 1/secretion in mycobacteria] to deliver virulence factors into host macrophages during infection. Despite its essential role in virulence, the mechanism of ESX-1 secretion is unclear. We found that the unstructured C terminus of the CFP-10 substrate was recognized by Rv3871, a cytosolic component of the ESX-1 system that itself interacts with the membrane protein Rv3870. Point mutations in the signal that abolished binding of CFP-10 to Rv3871 prevented secretion of the CFP-10 (culture filtrate protein, 10 kilodaltons)/ESAT-6 virulence factor complex. Attachment of the signal to yeast ubiquitin was sufficient for secretion from M. tuberculosis cells, demonstrating that this ESX-1 signal is portable.     

对推行兽药GMP工作的思考

对推行实施兽药GMP工作中存在的问题进行了分析,提出了建议和对策,并指出兽药行业的规范化管理,除了要推行实施兽药GMP外,还应加快对兽药的经营、使用、研发、临床试验等各环节的立法,建立健全机构,加强队伍建设,特别是要尽快推行兽药经营质量管理规范(GSP),否则兽药GMP将很难长期坚持下去.     

Development of a micro-sward technique for determining intake characteristics of perennial ryegrass varieties

Two diploid (Belramo and Glen), one tetraploid (Rosalin) and one tetraploid hybrid (Lolium× boucheanum Kunth) (AberExcel) intermediate-heading perennial ryegrass (Lolium perenne L.) varieties were sown in boxes (85×44×14 cm) to create micro-swards. Once established, these were defoliated at approximately 21-day intervals before being offered to individual yearling Simmental × Holstein beef heifers in short-term tests. Bite mass measured in these tests was compared with bite mass for groups of similar heifers which were rotationally stocked on the same four varieties. The boxes were weighed (± 0.1 g) before and after a period during which the heifers were allowed to take approximately 50 bites. The micro-sward methodology was developed in, 2002 and bite masses for Belramo, Glen, Rosalin and AberExcel, respectively, were 0.44, 0.46, 0.49 and 0.50 g DM bite^–1 compared with 0.48, 0.49, 0.50 and 0.50 g DM bite^–1 measured for rotationally stocked cattle. In 2003, the values for bite mass were 0.40, 0.41, 0.41 and 0.43 g DM bite^–1 measured using sward boxes and 0.31, 0.41, 0.34 and 0.37 g DM bite^–1 for rotationally stocked cattle. Whilst the rankings were similar within ploidy for the two assessment methods, there were no significant differences between the varieties. The micro-sward methodology provides a rapid, low-cost assessment method with a low seed requirement to screen for intake traits during breeding programmes and during variety evaluation. Now there is a need to develop further this methodology in order to allow small-scale simulation of other defoliation regimes, for example continuous stocking.     

Artificial Insemination of Swine: Fecundity of Boar Semen Stored in Beltsville TS (BTS), Modified Modena (MM), or MR-A and Inseminated on One,Three and Four Days After Collection12

Contents: A field trial was conducted on 1,463 farms in The Netherlands to compare the fertilizing capacity of boar spermatozoa extended in Beltsville TS (BTS), Modified (MM), or MR-A, and inseminated in 2896 sows and gilts on the first, third or fourth day following collection. Semen was collected, extended, and stored at 18°C at six different AI centers and inseminated by inseminators on their regular routes. Sows inseminated with BTS and MR-A extended semen had higher farrowing rates than MM (79.3, 77.6 us 50.4, P <.0001), higher total pigs per litter (11.4, 11.1 vs 10.0, P <.0001) and higher total pigs born alive (10.7, 10.5 vs 9.4, P <. 0001). The farrowing rate of gilts inseminated with BTS extended semen was superior to MM (73.5 vs. 50.2%, P <.004), while MR-A gave farrowing rates greater than MM (64.1 vs 50.2%, P =.06). There was no difference in litter data for gilts. Farrowing rates for 1 and 4 day semen were superior to 3 day semen (73.2, 73.8 vs 60.3, P <.0001). The semen inseminated on day 4 contained 6 billion sperm per dose rather than the 3 billion per dose for 1 and 3 day semen. Based on the results of this study, BTS and MR-A are effective diluents for extension and storage of boar semen for use within the same week of collection. In addition, semen extended in either BTS of MR-A and stored and inseminated on the fourth day after collection can give fertility equal to first day insemination if the sperm per insemination dose is doubled. Inhalt: Künstliche Besamung beim Schwein: Fruchtbarkeitsergebnisse von Ebersamen nach Flussigkonservierung mit BTS, MR-A oder modifiziertem Modena-Verdünner (MM) und Besamung am ersten, dritten und vierten Tag nach Samengewinnung In einem in den Niederlanden durchgeführten Feldversuch wurden 3 verschiedene Verdünner-Medien für Ebersamen in vivo verglichen. Die Medien waren Beltsville TS (BTS), modifizierter Modena-Verdünner (MM) und das spanische MR-A-Medium. Insgesamt wurden 2896 Jung- und Altsauen in 1463 Herden mit 1, 3 oder 4 Tage altem Sperma besamt. Der Versuch wurde als “split-sample”-Versuch an 6 Eberstationen durchgeführt, der verdunnte Samen bei + 18°Cgelagert und von den Besamungsbeauftragten im Rahmen ihrer regulären Fahrten eingesetzt. BTS- und MR-A-konserviertes Sperma gab bei Altsauen gegenüber MM höhere Abferkelraten (79,3, 77,6 vs 50,4%, P < 0.0001), eine höhere totale Wurfzahl (11,4, 11,1 vs 10,0, P < 0.0001) und eine höhere Anzahl lebendgeborener Ferkel)10,7, 10,5 vs 9,4, P < 0.0001). Die mit BTS-uerdünntem Samen inseminierten Jungsauen wiesen gegenüber MM höhere Abferkelraten auf(73.5 vs 50,2%, P < 0.004). MR-A-Sperma lag ebenfalls hüher als MM-Samen (641 vs 50,2%, P=0.06). Die Wurfgröβen der Jungsauen reigten keine Unterschiede. Die Abferkelergebnisse nach Besamung mit 1 und 4 Tage altem Samen lagen höher als die des 3 rage alten Spermas (73,2, 73,8 vs 60,3%; P < 0.0001). Die am Tag 4 verwendeten Samenportionen enthielten 6 Milliarden Spermien gegenüber 3 Milliarden der nach 1 bzw. 3 Tagen inseminierten Portionen. Die Ergebnkse dieses Versuches zeigen, daβ BTS und MR-A effektive Verdünnermedien sind und die Lagerung von Eberfrischsperma zum Einsatz innerhalb der Entnahmewoche ermoglichen. Darüber hinaus können mit BTS oder MR-A verdünntes und am Tag 4 nach Gewinnung eingesetrtes Sperma Fruchtbarkeitsergebnisse erzielt werden, die ebenso gut sind wie nach Verwendung des 1-Tage-Spermas, wenn die Spermienzahl pro Besamungsdosis verdoppelt wird.     

Characterization of Transgenic Pigs That Express Human Decay Accelerating Factor and Cell Membrane‐tethered Human Tissue Factor Pathway Inhibitor

The expression of human complement regulatory proteins (hCRP; hDAF, hCD59, and hMCP) in pig tissues has been suggested as one of strategies to overcome the hyperacute rejection (HAR) in pig‐to‐human transplantation. Expression of human tissue factor pathway inhibitor (hTFPI) in porcine endothelial cells has been suggested as a remedy to overcome microvascular thrombosis. To investigate the effects of these combined transgenes, we established transformed pig cells expressing human decay accelerating factor (hDAF) under the control of enhancer promoter (5′LTR‐PCMVIE), and the fusion protein (hTFPI/hCD4) consisting of the functional domains (K1 and K2) of hTFPI and membrane‐tethering domains (D3 and D4) of hCD4 under the control of PCMVIE. Transgenic pigs were generated with the transformed porcine cells through somatic cell nuclear transfer (SCNT) technology. Analysis of quantitative PCR and real‐time quantitative RT‐PCR showed that four copies of hDAF were integrated and 391 copies of hDAF mRNA expressed in the cells of the transgenic pig. The enhancing activity of 5′LTR was approximately 2 fold compared to CMVIE promoter only. The cell viability test showed that more than 80% of ear cells were viable in the presence of 50% human serum. The chromogenic substrate assay and immunocytochemical staining with tail cells showed that the TFPI activity of fusion protein was observed on the cell membrane. The membrane localization of hDAF and hTFPI proteins was observed by immunocytochemical staining, and the expression of transgenes in heart and liver tissues was also confirmed by immunohistochemistry.     

Triazole fungicides and the selection of resistance to medical triazoles in the opportunistic mould Aspergillus fumigatus

Azole resistance is an emerging problem in the opportunistic mould Aspergillus fumigatus. The triazoles are the most important agents for the management of Aspergillus diseases in humans. Selection for acquired resistance may occur in the hospital setting through exposure to high doses of azoles during azole therapy, but evidence is accumulating that A. fumigatus may become resistant to medical triazoles through environmental exposure to fungicides. The recovery of A. fumigatus isolates resistant to the medical triazoles from azole‐naive patients as well as from the environment strongly indicates an environmental route of resistance selection. Molecule alignment studies have identified five fungicides that share a very similar molecule structure with the medical triazoles, and thus may have selected for mechanisms that confer resistance to both groups of compounds. It is important to explore further the presumed fungicide‐driven route of resistance selection in order to implement effective preventive measures as the prevalence of azole resistance in A. fumigatus continues to increase and causes major challenges in the management of azole‐resistant Aspergillus diseases. Copyright © 2012 Society of Chemical Industry     

The 2001 epidemic of foot-and-mouth disease in the United Kingdom: epidemiological and meteorological case studies

The possibility of the airborne spread of foot-and-mouth disease during the 2001 epidemic in the uk has been investigated in three epidemiological case studies. On the basis of evidence from field investigations, and a simple meteorological analysis, it is concluded that the spread of disease was consistent with the airborne transport of virus. The distances ranged from less than 1 km to 16 km; six of the farms were over 6 km from the source and involved the passage of virus over the sea combined with meteorological conditions which strongly favoured airborne disease transmission. The results of detailed atmospheric modelling demonstrated that airborne virus could have challenged livestock on all the farms studied. However, with one exception the 24-hour average daily concentrations of the virus were significantly below the experimentally estimated threshold for infection. A detailed model intercomparison established that, under stable atmospheric conditions, peak concentrations of virus up to two orders of magnitude higher might have been experienced for short periods, owing to fluctuations within the plume of virus, and model limitations. This finding would significantly reduce the apparent discrepancy between the experimentally estimated threshold for infection and the modelling results.